The goal of these studies is to probe the mechanism by which glucocorticoids initiate DNA synthesis and cell division in quiescent cultured fibroblasts. Most of the studies will be conducted with 3T3 mouse fibroblasts, although some experiments will be performed on human diploid foreskin fibroblasts. The response of 3T3 cells to added glucocorticoids has been somewhat variable, and we will continue our efforts to identify parameters which influence the ability of glucocorticoids to initiate proliferation in these cells. Our studies have demonstrated that serum is required for the initiation, and preliminary experiments indicate that there are serum components required for the response that either are labile and decay spontaneously at 37 degrees, or are rapidly depleted by 3T3 cells. Another parameter which appears to affect the responsiveness of quiescent 3T3 cells to added glucocorticoids is the density at which the cells are seeded. As soon as the response to glucocorticoids is optimized, we will proceed with our plans to isolate 3T3 cells that are unresponsive to glucocorticoid addition, and determine if these cells have alterations in glucocorticoid specific cell components (e.g., cytoplasmic glucocorticoid receptor) and interactions among these components. These studies will allow us to determine if these components are causally involved in the initiation of proliferation by glucocorticoids. BIBLIOGRAPHIC REFERENCES: Growth Limitation of 3T3 Mouse Fibroblasts by Available Growth Surface Area and Medium Components. Cornelia R. Thrash and Dennis D. Cunningham. Journal of Cellular Physiology, 86 301 (1975). Effects of Added Proteases on Concanavalin A-Specific Agglutinability and Proliferation of Quiescent Fibroblasts. Dennis D. Cunningham and Tsung-Shang Ho. In Proteases and Biological Control. Published by Cold Spring Harbor Laboratory. 1975. pp. 795.